The overall, long-term objective of this project is elucidation of the biochemical mechanisms underlying granulocyte proliferation and differentiation. Our approach centers around development of convenient and quantitative radioimmunoassays (RIA), immunocytofluorescence (IF) procedures and enzymatic assays for studying the in vitro synthesis of granulocyte-specific proteins in a readily accessible animal model, the mouse. In particular, we are studying the proteins myeloperoxidase (MPO) and lactoferrin (LF). Each is synthesized at a different period of the granulocyte maturation sequence, and consequently, each is initially associated with a different recognizable cell type (promyelocyte and myelocyte, respectively). We have developed a density-gradient fractionation method for the purification of PMN from mouse peritoneal exudate cells (PEC). It was observed that PMN showed a bimodal distribution into high density (HD, 1.09gm/ml) and low density (LD, 1.06 gm/ml) fractions. The HD fraction progressively shifted to the LD position as the time following injection of stimulator increased (1% glycogen, 2-9 hr). Qualitative changes (iodinating and decarboxylating activity, inhibition by aminotriazole) were observed in MPO activity (guaiacol oxidation) between the HD and LD cells. Rabbit antisera prepared against MPO purified from bone marrow cells did not distinguish between PMN and monocytes/macrophages by IF. These observations support the notion that the two activities are the same, and that reported differences may be due to changes in enzyme activity related to the functional state of the cell.